The brain becomes infected within days after peripheral HIV infection and viral presence persists despite cART. CNS HIV infection often results in the development of HIV Associated Neurocognitive Disorders, or HAND, that are mediated, at least in part, by ongoing neuroinflammation and low level viral persistence that cause CNS damage. A major mechanism by which virus enters the brain is by the transmigration of HIV-infected monocytes and perhaps T cells across the BBB. Under non-pathological conditions, the transmigration of leukocytes across the CNS vasculature does not disrupt the BBB because specific interactions between adhesion molecules, adherens junction and tight junction proteins (termed "cell junction proteins") on leukocytes and BBB cells maintain the impermeability of these vessels during leukocyte diapedesis. However, during the pathogenesis of NeuroAIDS, leukocyte infiltration into the CNS is associated with BBB compromise. Thus the molecular interactions inherent in leukocyte diapedesis across the BBB are altered, resulting in increased BBB disruption. Our data indicate that HIV infection of monocytes and T cells plus CCL2 as a chemoattractant, but not other chemokines, cause increased transmigration of these cells across a tissue culture model of the BBB and disruption of that barrier. We demonstrated that the virus as well CCL2 induced profound changes in expression, processing and function of diverse cell junction proteins on both leukocytes and brain microvascular endothelial cells. We hypothesize that HIV infection of monocytes and exposure to CCL2 alter their expression, localization, phosphorylation and shedding of cell junction proteins and that these alterations result in aberrant leukocyte/BBB cell interactions, leading to increased transmigration and barrier disruption, contributing to the neuroinflammation and viral damage that characterize NeuroAIDS This renewal is to continue our studies of the mechanisms of transmigration of HIV infected cells across the BBB in response to CCL2, with emphasis on a specific monocyte population that we showed preferentially transmigrates across the BBB in response to CCL2 and is highly susceptible to HIV infection. We will focus on the cell junction molecules PECAM-1, CD99, JAM-A, and ALCAM, as critical proteins involved in dysregulation of monocyte transmigration and disruption of BBB integrity. We also identified additional proteins that may facilitate dysregulated transmigration as well as monocyte susceptibility to HIV infection, and we will examine their expression and function on monocytes. We will examine the contribution of intact and soluble junction proteins to increased transmigration, and determine the route, paracellular or transcellular, of transmigration of uninfected and HIV monocytes. We will demonstrate the in vivo significance of these processes using an SIV model. PUBLIC HEALTH RELEVANCE: HIV infection and HIV associated neurocognitive disorders (HAND) are major emerging global health problems. The prevalence of HAND is increasing as people with AIDS are living longer due to success of antiretroviral therapies (ART). HAND is mediated, at least in part, by inflammation and viral persistence within the CNS, which are present even in many individuals on ART. Our group demonstrated that HIV infected leukocytes transmigrate more exuberantly across the BBB in response to CCL2 as chemoattractant, but not to other chemokines. We propose that CCL2 and/or HIV infection result in profound changes in expression, localization and shedding of the cell junction proteins ALCAM, PECAM-1, JAM- A and CD99 on monocytes and brain microvascular endothelial cells (BMVEC) that enable infected cells to have such increased transmigration across the BBB. In this resubmitted competing renewal we will determine the mechanisms by which these junction proteins are dysregulated on monocytes and BMVEC to result in BBB disruption and increased transmigration in response to CCL2. We propose that targeting these cell junction proteins may be a novel interventional strategy to reduce HIV CNS impairment.